Transglutaminase-2 (TGase-2) is a GTP-binding protein/protein crosslinking enzyme implicated in various pathological conditions including coeliac disease, neurodegenerative disorders, and aggressive cancers. The mechanistic basis by which TGase-2 contributes to many of these disorders is poorly understood. Recently, we obtained exciting new clues regarding the roles of TGase-2 in human cancers by identifying it as a unique EGF receptor (EGFR)-signaling partner. Specifically, we found TGase-2 enhances EGFR expression by protecting EGFRs against the actions of c-Cbl, as well as functions as a novel scaffold to mediate PI-3K activation. TGase-2 expression is inversely correlated with brain cancer survival, and it is specifically expressed together with an oncogenic variant of the EGFR, the EGFRvIII, in a highly aggressive subset of glioma initiator (stem) cells (GSCs), called mesenchymal (MES) GSCs. Moreover, TGase-2 and EGFRvIII are among the cargo of microvesicles (MVs) shed by MES GSCs, offering a potential mechanism by which MES GSCs transfer these important signaling proteins to surrounding cells to reprogram their microenvironment. In this proposal, we will build on these discoveries to establish the mechanisms by which TGase-2 contributes to disease states. We will focus on its role in glioblastoma (GBM) through the combined efforts of two laboratories with complimentary expertise in EGFR-signaling (Cerione) and translational studies of GSCs (Nakano), as follows: 1) Test the hypothesis that TGase-2 plays novel roles in the actions of the oncogenic EGFRvIII and other receptors implicated in GBM. Based on our recent findings, we will determine whether TGase-2 adversely affects patient survival by protecting the EGFRvIII, as well as other receptors linked to GBM, against c-Cbl-catalyzed degradation and/or by enhancing their signaling to PI-3K/Akt/mTORC1. 2) Test the hypothesis that TGase-2 and the EGFRvIII play an essential part in the intercellular signaling mediated by MVs from MES GSCs. We will determine whether this extremely aggressive sub-set of GSCs use MVs to `communicate' with less aggressive GSCs or with differentiated cancer cells to re-program them for tumor formation. 3) Test the idea that targeting TGase-2 through different strategies blocks the actions of MES GSCs in mouse models for GBM. The expectation is these studies will be broadly relevant toward understanding the biological roles of TGase-2 in disease, as well as identify novel therapeutic targets for EGFR/EGFRvIII-driven cancers such as GBM.